Korean Non-small Cell Lung Cancer Research Articles

Proteogenomic analysis reveals non-small cell lung cancer subtypes predicting chromosome instability, and tumor microenvironment

Non-small cell lung cancer (NSCLC) is histologically classified into lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LSCC). However, some tumors are histologically ambiguous and other pathophysiological features or microenvironmental factors may be more prominent. Here we report integrative multiomics analyses using data for 229 patients from a Korean NSCLC cohort and 462 patients from previous multiomics studies. Histological examination reveals five molecular subtypes, one of which is a NSCLC subtype with PI3K-Akt pathway upregulation, showing a high proportion of metastasis and poor survival outcomes regardless of any specific NSCLC histology. Proliferative subtypes are present in LUAD and LSCC, which show strong associations with whole genome doubling (WGD) events. Comprehensive characterization of the immune microenvironment reveals various immune cell compositions and neoantigen loads across molecular subtypes, which predicting different prognoses. Immunological subtypes exhibit a hot tumor-enriched state and a higher efficacy of adjuvant therapy.

P1.23-05 Clinical Utility of Plasma-Based Comprehensive Genomic Profiling in Korean Non-small Cell Lung Cancer Patients

P1.23-05 Clinical Utility of Plasma-Based Comprehensive Genomic Profiling in Korean Non-small Cell Lung Cancer Patients

Psychometric Validation of the Korean Version of the Cancer Survivors' Unmet Needs (CaSUN) Scale among Korean Non-Small Cell Lung Cancer Survivors.

The purpose of the study was to validate the Korean version of Cancer Survivors' Unmet Needs (CaSUN) scale among non-small cell lung cancer survivors. Participants were recruited from outpatient clinics at the Samsung Medical Center in Seoul, South Korea, from January to October 2020. Participants completed a survey questionnaire that included the CaSUN. Exploratory and confirmatory factor analysis and Pearson's correlations were used to evaluate the reliability and validity of the Korean version of the CaSUN (CaSUN-K). We also tested known-group validity using an independent t test or ANOVA. In total, 949 provided informed consent and all of which completed the questionnaire. Among the 949 patients, 529 (55.7%) were male; the mean age and median time since the end of active treatment (standard deviation) was 63.4±8.8 years and the median was 18 months. Although the factor loadings were different from those for the original scale, the Cronbach's alpha coefficients of the six domains in the CaSUN-K ranged from 0.68 to 0.95, indicating satisfactory internal consistency. In the CFA, the goodness-of-fit indices for the CaSUN-K were high. Moderate correlations demonstrated the convergent validity of CaSUN-K with the relevant questionnaire. More than 60% of the participants reported information-related unmet needs, and the CaSUN-K discriminated between the needs reported by the different subgroups that we analyzed. The CaSUN-K is a reliable and valid measure for assessing the unmet needs in a cancer population, thus this tool help population to receive timely, targeted, and relevant care.

Abstract 2343: The incidence of BRAF mutation in non-small cell lung cancers is low in Korean patients

Abstract Introduction: BRAF mutation is present in about 8% of all cancer. Oncogenic activity of BRAF V600E mutation was established in lung cancers. Dabrafenib and trametinib combination therapy is preferred first line therapy for advanced or metastatic lung cancer with BRAF V600E mutation. However, there is only little basic knowledge of BRAF mutational status among Korean population. In this study, BRAF mutation, particularly BRAF V600E mutational status was explored among Korean non-small cell lung cancer patients. The clinical and pathologic characteristics of BRAF V600E mutation in non-small cell lung cancers were also investigated. Materials and Methods: The final cohort was 378 cases of primary non-small cell lung cancers. Fromalin-fixed paraffin embedded (FFPE) tissue blocks were used. PNA-mediated clamping PCR for BRAF V600, BRAF V600E real-time PCR and immunohistochemistry for mutation specific monoclonal antibody of Ventana VE1 were performed on all 378 cases of non-small cell lung cancers. For positive cases, direct sequencing was performed for validation. Results: BRAF V600 mutation was detected in five cases using PNA clamping method among 378 non-small cell carcinoma cases. By using BRAF real-time PCR, a BRAF V600E mutation was detected in 3 patients among total 378 cohort. There were 2 discordant cases between PNA clamping and real-time PCR. On immunohistochemistry, 3 patients showed positive staining for tumor cytoplasm and all positive cases also had positive results in real-time PCR. Direct Sanger sequencing was done in 5 cases that had any positive results for 3 methods. The results of Sanger sequencing was the same with those of real-time PCR and immunohistochemistry. Direct sequencing using PAN PCR product was performed on 2 negative cases for validation. On sequencing using PNA clamping PCR product, all cases showed mutation for BRAF gene other than V600E genotype. Finally, there were 3 cases (0.8%) of BRAF V600E mutation and 2 cases of BRAF mutation (0.5%) other than V600E genotype. Among BRAF mutated patients, there were 2 never smokers and 3 ever smokers. All patients harboring BRAF mutation had no concomitant EGFR or ALK aberrations. Conclusion: BRAF mutation incidence is relatively lower than that of Western data set and similar to that of Japanese report. Immunohistochemistry for Ventana VE1 antibody can be a useful screening method to detect BRAF V600E mutation. Citation Format: Ahrong Kim, Hyo Yeong Ahn, Chang Hun Lee, Young Dae Kim, Min Ki Lee, Yeon Joo Jeong, So Jeong Lee. The incidence of BRAF mutation in non-small cell lung cancers is low in Korean patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2343.

Abstract B16: High-resolution metagenomic profiling revealed gut microbiome features for the response to cancer immunotherapy reproducible among multiple cohorts of patients

Abstract Cancer immunotherapy is currently standard of treatment showing significant clinical benefits to cancer patients, but the beneficial effects are limited to a relatively small subset of patients. Of various factors that may affect the efficacy of cancer immunotherapy, the gut microbiome is receiving more attention because of its strong correlation with immune regulation. Recently, independent studies based on Western populations have identified several taxa that are differentially abundant between responders and nonresponders. However, the reported taxa associated with anti-PD-1 response differ entirely among these studies. In this study, we performed 1) deep (>100M reads in average) metagenome shotgun sequencing on the fecal sample of 49 Korean non-small cell lung cancer (NSCLC) patients, 2) taxonomic profiling using a highly comprehensive gut microbiome-specific genome database, and 3) gene-level profiling with the gut microbiome specific gene catalog to improve the resolution of metagenomic analysis. With the high-resolution metagenomic profiling, we found several taxonomic and functional signatures associated with the response of anti-PD-1 therapy. Interestingly, we found that some of the signatures were reproduced in five independent datasets regardless of the cancer types (melanoma, NSCLC, and renal cell carcinoma) and the type of immunotherapy (anti-PD-1, anti-CTLA4, and the combination of them). For instance, a responder-enriched taxon in the Korean cohort was more frequently detected in the responders in all cohorts (abundance fold-change > 2) and significantly enriched (p<0.05) in two cohorts. Moreover, when we performed an association analysis with the combined profile, the taxon was identified with the highest significance among the differentially abundant taxa (fold-change=2.41, p=2.5e-06). Consistent with the result of the taxon-level analysis, we found that the genes encoded by the taxon were also enriched in responders in multiple studies. In summary, using high-resolution metagenomic profiling, we identified gut microbiome features for the response to cancer immunotherapy shared across multiple cohorts for the first time, to the best of our knowledge. Citation Format: Chan Yeong Kim, Beung-Chul Ahn, Min Hee Hong, Yoo Jin Chun, Hye Ryun Kim, Insuk Lee. High-resolution metagenomic profiling revealed gut microbiome features for the response to cancer immunotherapy reproducible among multiple cohorts of patients [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr B16.

Characteristics and outcomes of RET-rearranged Korean non-small cell lung cancer patients in real-world practice.

Since the first discovery of rearranged during transfection (RET) fusion in lung adenocarcinoma in 2011, two tyrosine kinase inhibitors, namely vandetanib and cabozantinib, are currently available. Despite favorable outcomes in systemic control, the intracranial therapeutic response remains insufficient. In this study, the clinical characteristics and outcomes of non-small cell lung cancer (NSCLC) patients with RET rearrangements were analyzed. Patients with NSCLC harboring RET fusion who received treatment between January 2006 and January 2018 were analyzed. RET rearrangement was identified by FISH or NGS. A total of 59 patients were identified. About half of the patients were female (47.5%) and never smokers (50.9%). Most patients had adenocarcinoma (89.8%). A total of 17 patients (28.8%) had an intracranial lesion at the initial diagnosis of stage IV disease, and 11 additional patients (18.6%) developed intracranial metastases during follow-up. The median time to development of intracranial metastases was 19.0months (95% CI: 9.6-28.5), resulting in a >60% cumulative incidence of brain metastasis at 24months. The systemic efficacy of pemetrexed-based regimens was favorable with progression-free survival of 9.0 (95% CI: 6.9-11.2) and OS of 24.1 (95% CI: 15.2-33.0) months. The median progression-free survival for vandetanib and immunotherapy was 2.9 (95% CI: 2.0-3.8) and 2.1 (95% CI: 1.6-2.6) months, respectively. Given the likelihood of RET-rearranged NSCLC progressing to intracranial metastases and the absence of apparent clinical benefit of currently available targeted or immunotherapeutic agents, development of novel treatment with higher selectivity and better penetration of the blood-brain barrier remains a priority.

Characteristics and Clinical Outcomes of Non-small Cell Lung Cancer Patients in Korea With MET Exon 14 Skipping.

MET exon 14 skipping occurs in 3-4% of patients with lung adenocarcinomas. In this study, we performed a comprehensive analysis of clinical data from Korean non-small cell lung cancer (NSCLC) patients with MET exon 14 skipping. Overall, 1,020 patients diagnosed with NSCLC between January 2015 and July 2017 were analyzed by next-generation sequencing. MET exon 14 skipping was identified in 20 NSCLC patients (1.9%). The median age was 69 years (range=39-86 years), 60.0% were male, and most (55.0%) were ever-smokers. For first-line chemotherapy, the median overall survival was 9.5 months and progression-free survival was 4.0 months, respectively. Twelve patients received pemetrexed-based chemotherapy and the overall response rate was 33.3% (4/12). Among four crizotinib-treated patients, one continued therapy for 8 months with the best response being disease stability. Given the poor clinical outcome and response to therapy for NSCLC, and the availability of promising anti-tumor MET inhibitors, screening for the MET exon 14 skip mutation should be incorporated into clinical practice.

P1.16 A Study on Clinicopathological Features and Diagnostic Methods of ALK Fusion-Positive Non-Small Cell Lung Cancer in KR

Lung cancer is one of the most common malignancies and the leading cause of cancer-related death in Korea. A lot of efforts have been put into to develop new, more effective treatments and biological markers to predict the therapeutic responses, leading to the discovery of various genetic changes, the so-called “growth drivers” of carcinogenesis. Some genetic alterations have become the new treatment targets, and there have been suggestions that different mutations are associated with different clinicopathological characteristics and prognosis. In this study we aimed to evaluate the status of the key “driver” mutation ALK fusion in Korean non-small cell lung cancer patients and its association with the clinicopathological characteristics, including the presence of other genetic mutations. We also compared different methods for ALK fusion detection, i.e., fluorescence in situ hybridization (FISH), immunohistochemistry (IHC), and next-generation sequencing (NGS) analyses to find out which method would the most effective. A total of 482 NSCLC patients who underwent ALK FISH test were evaluated for clinicopathological features such as age, sex, smoking history, tumor stage, histological subtype, immunohistochemical profile including ALK, EGFR mutation status and survival. Some ALK FISH-positive and -negative cancers were newly submitted to NGS analysis for DNA and RNA alterations. The ALK fusion-positive tumors were associated with younger age, more female patients, frequent nodal metastases and advanced stage, and short survival. The diagnostic agreement rates among ALK FISH, ALK IHC, and NGS ranged from about 60 to 90% and when considering the patient outcome as a measurement of the test efficacy, the ALK FISH may not be the most reliable one in diagnosis of ALK fusion-positive lung cancer. The NGS analysis detected various additional genetic alterations never considered at the time of diagnosis, including NPM3-TRK rearrangement. Comparing the results of ALK FISH, IHC and NGS, we concluded in practice ALK testing should better be diversified concerning FISH and IHC, and NGS analysis would be a good alternative of ALK FISH with an additional advantage of simultaneous detection of different mutations.

Potential Oncogenic Role of the Papillary Renal Cell Carcinoma Gene in Non-Small Cell Lung Cancers.

PurposePapillary renal cell carcinoma (PRCC) gene, which located in 1q23.1, is recurrently amplified in non-small cell lung cancer (NSCLC). However, it is unknown whether PRCC is overexpressed in primary NSCLCs and whether PRCC overexpression contributes to lung tumorigenesis. In this study, we aimed to identify the profiles of PRCC expression in Korean NSCLC patients and to elucidate the role of PRCC overexpression on lung tumorigenesis.Materials and MethodsWe performed immunohistochemistry analysis with a tissue array containing 161 primary NSCLCs. Small interfering RNA targeting PRCC (siPRCC) was transfected into two lung cancer cell lines (NCI-H358 and A549), after which tumor growth, migration, and invasion were observed. Expressions of cell proliferation-, cell cycle-, and metastasis-related molecules were examined by Western blot analysis. We also explored the in vivo effect of PRCC silencing.ResultsPRCC overexpression was recurrently observed in NSCLCs (95/161, 59%). After siPRCC treatment, tumor cell proliferation, colony formation, and anchorage independent growth were significantly reduced (p<0.001 for all three effects). Migration and invasiveness were also significantly repressed (p<0.001 for both effects). Reflecting cell proliferation, cell cycle, and metastasis, the expressions of Ki67, cyclin D1, AKT-1, pAKT, NF-kB p65, vimentin and CXCL-12 were found to be downregulated. Through mouse xenograft analysis, we confirmed that PRCC silencing significantly repressed a xenograft tumor mass in vivo (p<0.001).ConclusionThe present data provide evidence that PRCC overexpression is involved in the tumorigenesis and progression of lung cancer.

A pilot study identifying a potential plasma biomarker for determining EGFR mutations in exons19 or 21 in lung cancer patients.

The most common type of lung cancer is non-small cell lung cancer (NSCLC), which is frequently characterized by a mutation in the epidermal growth factor receptor (EGFR). Determining the presence of an EGFR mutation in lung cancer is important, as it determines the type of treatment that a patients will receive. Therefore, the aim of the present study was to apply high-resolution metabolomics (HRM) using liquid chromatography-mass spectrometry to identify significant compounds in human plasma samples obtained from South Korean NSCLC patients, as potential biomarkers for providing early detection and diagnosis of minimally-invasive NSCLC. The metabolic differences between lung cancer patients without EGFR mutations were compared with patients harboring EGFR mutations. Univariate analysis was performed, with a false discovery rate of q=0.05, in order to identify significant metabolites between the two groups. In addition, hierarchical clustering analysis was performed to discriminate between the metabolic profiles of the two groups. Furthermore, the significant metabolites were identified and mapped using Mummichog software, in order to generate a potential metabolic network model. Using metabolome-wide association studies, metabolic alterations were identified. Linoleic acid [303.23 m/z, (M+Na)+], 5-methyl tetrahydrofolate [231.10 m/z, (M+2H)+] and N-succinyl-L-glutamate-5 semialdehyde [254.06 m/z, (M+Na)+], were observed to be elevated in patients harboring EGFR mutations, whereas tetradecanoyl carnitine [394.29 m/z, (M+Na)+] was observed to be reduced. This suggests that these compounds may be affected by the EGFR mutation. In conclusion, the present study identified four potential biomarkers in patients with EGFR mutations, using HRM combined with pathway analysis. These results may facilitate the development of novel diagnostic tools for EGFR mutation detection in patients with lung cancer.

Gefitinib-Induced Interstitial Lung Disease in Korean Lung Cancer Patients.

PurposeInterstitial lung disease (ILD) is a serious adverse effect of gefitinib. We examined the incidence and clinical characteristics of drug-induced ILD in Korean non-small cell lung carcinoma patients treated with gefitinib.Materials and MethodsA retrospective cohort study was performed in non-small cell lung cancer (NSCLC) patients who started gefitinib treatment at Seoul National University Hospital from January 2002 through December 2011. Patients who developed new abnormal radiologic findings with respiratory symptoms after gefitinib treatment were defined as having possible adverse pulmonary reactions. The patients’ medical records were reviewed independently by investigators to identify the causes of pulmonary toxicities.ResultsAmong the 1,114 patients evaluated, 128 patients (11.5%) developed pulmonary adverse reactions after taking gefitinib. An infectious complication occurred in 98 patients (8.8%) and 15 patients (1.3%) developed ILD. Nine of the 15 patients (60.0%) with gefitinib-induced ILD experienced a fatal clinical course that met either the Common Terminology Criteria for Adverse Events grade 4 (n=3) or grade 5 (n=6). In the multivariate analysis, a lower serum albumin level (≤ 3.0 g/dL) at baseline was significantly associated with the development of gefitinib-induced ILD (odds ratio, 3.91; 95% confidence interval, 1.20 to 12.71).ConclusionThe incidence of gefitinib-induced ILD in Korean NSCLC patients was similar to that reported worldwide, but lower than values reported for Japanese population. ILD was usually a life-threatening adverse effect of gefitinib, and the development of ILD was significantly associated with a lower baseline serum albumin level.

Pathway-Based Analysis Using Genome-wide Association Data from a Korean Non-Small Cell Lung Cancer Study

Pathway-based analysis, used in conjunction with genome-wide association study (GWAS) techniques, is a powerful tool to detect subtle but systematic patterns in genome that can help elucidate complex diseases, like cancers. Here, we stepped back from genetic polymorphisms at a single locus and examined how multiple association signals can be orchestrated to find pathways related to lung cancer susceptibility. We used single-nucleotide polymorphism (SNP) array data from 869 non-small cell lung cancer (NSCLC) cases from a previous GWAS at the National Cancer Center and 1,533 controls from the Korean Association Resource project for the pathway-based analysis. After mapping single-nucleotide polymorphisms to genes, considering their coding region and regulatory elements (±20 kbp), multivariate logistic regression of additive and dominant genetic models were fitted against disease status, with adjustments for age, gender, and smoking status. Pathway statistics were evaluated using Gene Set Enrichment Analysis (GSEA) and Adaptive Rank Truncated Product (ARTP) methods. Among 880 pathways, 11 showed relatively significant statistics compared to our positive controls (PGSEA≤0.025, false discovery rate≤0.25). Candidate pathways were validated using the ARTP method and similarities between pathways were computed against each other. The top-ranked pathways were ABC Transporters (PGSEA<0.001, PARTP = 0.001), VEGF Signaling Pathway (PGSEA<0.001, PARTP = 0.008), G1/S Check Point (PGSEA = 0.004, PARTP = 0.013), and NRAGE Signals Death through JNK (PGSEA = 0.006, PARTP = 0.001). Our results demonstrate that pathway analysis can shed light on post-GWAS research and help identify potential targets for cancer susceptibility.

Abstract 4461: Poor predictive power of BIM deletion polymorphism for EGFR tyrosine kinase inhibitor outcome in non-small cell lung cancer patients harboring EGFR mutation.

Abstract Background. Recently, a germline deletion polymorphism of BIM was reported to predict tyrosine kinase inhibitor (TKI) outcome in patients with chronic myelogenous leukemia or advanced non-small cell lung cancer (NSCLC) harboring epidermal growth factor receptor (EGFR) mutation. In this study, we examined the predictive role of BIM deletion in a Korean NSCLC cohort. Also, we explored genetic alteration profile of patients exhibiting primary resistance to EGFR TKI treatment despite EGFR mutation. Methods. Data from 197 consecutive patients who had stage IIIB/IV NSCLC with TKI-sensitive EGFR mutation were collected from the NSCLC database of Seoul National University Hospital between 2006 and 2011. BIM deletion polymorphism was genotyped with polymerase-chain reaction (PCR) and visualized with 2% agarose gel electrophoresis. Additionally, we also examined putative clinical predictors of EGFR TKI outcome, including line of treatment, EGFR genotype, and smoking status. For patients who exhibited primary resistance to EGFR TKI treatment, we performed additional molecular analysis including fluorescence in situ hybridization for MET amplification and ALK fusion, and targeted deep sequencing of 46 cancer-related genes to explore the mechanism of primary resistance. Results. In this cohort, the median progression-free survival (PFS) for patients receiving EGFR TKIs was 11.9 months, and the objective response rate was 78.8%. Patients with wild-type BIM (n = 172; 89.1%) and BIM deletion (n = 21; 10.9%) exhibited no difference in median PFS after EGFR TKI treatment (11.3 months for wild-type BIM, 11.9 months for BIM deletion; P = 0.791). Also, the line of treatment, the genotype of the EGFR-activating mutation, and smoking were not predictive of PFS for EGFR TKIs. However, patients with recurrent disease after curative surgical resection had a longer PFS than patients with initial stage IV disease (16.0 months for recurrent disease, 10.0 months for stage IV disease at initial presentation; P = 0.007). From the examination of pretreatment tissue of 11 patients exhibiting primary resistance, de novo EGFR T790M mutation was identified in one patient. We also found a patient with de novo MET amplification and another patient with ALK fusion. Targeted deep sequencing identified no recurrent, coexistent drivers of NSCLC. Conclusion. BIM deletion polymorphism showed poor predictive role in EGFR-mutant NSCLC patient cohort, although their initial report indicated remarkable predictive power. In a small portion of cases with primary resistance, we identified coexistent genetic alterations of cancer-related genes that could explain primary resistance. Our result suggests that the mechanism of primary resistance might be heterogeneous. Citation Format: June Koo Lee, Jong-Yeon Shin, Soyeon Kim, Sunhwa Lee, Changho Park, Ji-Yeon Kim, Youngil Koh, Bhumsuk Keam, Hye Sook Min, Tae Min Kim, Yoon-Kyung Jeon, Dong-Wan Kim, Doo Hyun Chung, Dae Seog Heo, Se-Hoon Lee, Jong-il Kim. Poor predictive power of BIM deletion polymorphism for EGFR tyrosine kinase inhibitor outcome in non-small cell lung cancer patients harboring EGFR mutation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4461. doi:10.1158/1538-7445.AM2013-4461

RET Fusion Genes in Korean Non-Small Cell Lung Cancer

Recently, rearranged during transfection (RET) fusions have been identified in approximately 1% of non-small cell lung cancer (NSCLC). To know the prevalence of RET fusion genes in Korean NSCLCs, we examined the RET fusion genes in 156 surgically resected NSCLCs using a reverse transcriptase polymerase chain reaction. Two KIF5B-RET fusions and one CCDC6-RET fusion were identified. All three patients were females and never smokers with adenocarcinomas. RET fusion genes were mutually exclusive from EGFR, KRAS mutations and EML4-ALK fusion. RET fusion genes occur 1.9% (3 of 156) of surgically treated NSCLC patients in Koreans.

Quantitative analysis of multiple gene promoter methylation in Korean non-small cell lung cancer patients and its association study with cancer risk factor and survival

The methylation profiles of nine cancer-associated genes were compared with the percent of methylation ratio (PMR) using quantitative methylation RT-PCR in 16 normal lung tissue samples and 80 non-small cell lung cancer (NSCLC) tissues samples. The methylation of nine cancer-associated genes showed a significant increase in tumors compared to matched normal lung tissues, and the level of gene methylation in NSCLC patients containing both adjacent nontumor and tumor tissues showed a significant increase compared to normal lung tissues (P<0.05). The hypermethylation of the p16, RASSF1A, and CYP1B1 genes were correlated with age at diagnosis. APC, RASSF1A, and CYP1B1gene hypermethylation was correlated with smoking status according to a cut-off of PMR values of 10. TWIST1 gene hypermethylation was correlated with histologic types. Kaplan-Meier survival analysis using the methylation status of nine genes demonstrated that only p16 gene hypermethylation was associated with the survival period. Our results of this study provide evidence that hypermethylation of cancer-associated genes containing a tumor suppressor may serve as an early detection marker for lung cancer progression.

High Incidence of EGFR Mutations in Korean Men Smokers with No Intratumoral Heterogeneity of Lung Adenocarcinomas: Correlation with Histologic Subtypes, EGFR/TTF-1 Expressions, and Clinical Features

Epidermal growth factor receptor (EGFR) mutation has been known to be associated with adenocarcinoma with bronchioloalveolar carcinoma (BAC; lepidic) feature. This study was aimed to characterize the frequency of EGFR mutations and their association with histologic subtypes in Korean nonsmall cell lung cancer (NSCLC) patients. Three hundred eighty-two (88 biopsies and 294 resections) NSCLC patients were investigated for EGFR mutations (exons 18-21) by polymerase chain reaction and direct sequencing method. For the resected adenocarcinoma specimens, histologic subtypes were classified according to both 2004 World Health Organization classification and 2011 International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society classification. The results were correlated with EGFR mutation and clinicopathologic features. EGFR mutations were detected in 196 of 382 NSCLCs (51.3%) and were more frequent in women than in men (65.7% versus 34.3%, p < 0.001) and in nonsmokers than in smokers (63.4% versus 32.0%, p < 0.001). Regarding histologic subtypes of adenocarcinoma, mixed acinar and BAC pattern showed the most frequent EGFR mutation (67.6%), followed by mixed papillary and acinar (65.2%), mixed solid and acinar (38.2%), micropapillary and acinar (30.4%), and acinar and mucinous BAC (13.3%). In addition, EGFR mutations were more frequently observed in tumors with BAC or papillary components than those with mucinous BAC or solid components. Identical EGFR mutations were detected in a single tumor showing mixed histological features. EGFR protein expression was seen more frequently in tumors with EGFR mutations than those without EGFR mutations (75.3% versus 24.7%, p=0.003). EGFR mutations were significantly more common in tumors with thyroid transcription factor-1 (TTF-1) expression than those without TTF-1 (p < 0.001), and almost all (92.7%) mutated adenocarcinomas were TTF-1 positive. The incidence of EGFR mutations is variable according to histologic subtypes, gender, and smoking history. The mixed acinar and BAC and papillary and acinar subtypes, the presence of BAC (lepidic) or papillary components, EGFR, and TTF-1 protein expression can predict higher EGFR mutation in lung adenocarcinoma. However, intratumoral heterogeneity of EGFR mutation was not found. In addition, relatively high incidence of EGFR mutations in Korean men who smoked with adenocarcinoma histology suggests that these patients should not be left behind EGFR mutation test.

The Author Response: EML4-ALK Fusion Gene in Korean Non-Small Cell Lung Cancer

I would like to thank the interest and comments on our paper entitled fusion gene in Korean non-small cell lung (1). In this study, we examined EML4-ALK fusion variants in Korean non-small cell lung cancers (NSCLCs) via reverse-transcriptase-polymerase chain reaction (RT-PCR) using primers designed to detect EML4-ALK fusion variants (variants 1, 2, 3a, 3b, 4, 5a, 5b, 6, and 7) that have been previously identified (2, 3). Our study demonstrated the spectrum and frequency of EML4-ALK fusion variants in Korean NSCLCs, which were different from those in other ethnic populations. I agree with the comment that the RT-PCR technology for identification of ALK fusion variants has several limitations. As pointed out in this comment, there are multiple EML4-ALK fusion variants and non-EML4 fusion partners, such as KIF5B, and KLC1 (2-5); therefore, any PCR-based strategy must incorporate validated primer pairs for all known ALK fusions. Another limitation is that given that most specimens from lung cancer patients are stored as formalin-fixed paraffin embedded tissue, the RNA may have been substantially degraded relative to non-fixed, fresh-frozen tissue. In addition, it has been reported that PCR-based detection of EML4-ALK can yield positive results in the absence of detectable ALK-rearrangement in both tumor and non-tumor tissues, suggesting a propensity for false positive results (6). The aim of our study was to examine the profile of known EML4-ALK fusion variants in Korean NSCLCs and was not to detect the presence of ALK fusion to other gene partners. The limitations of the RT-PCR analysis, such as the necessity of available high-quality RNA and the propensity of false positive results, were briefly discussed in the paper. I agree with the suggestion that long-distance-PCR or long distance inverse-PCR could be used to identify all EML4-ALK fusion variants as well as other ALK rearrangements having known or even unknown fusion partner genes (7, 8).

The different efficacy of gefitinib or erlotinib according to epidermal growth factor receptor exon 19 and exon 21 mutations in Korean non-small cell lung cancer patients

Epidermal growth factor receptor (EGFR) mutations are associated with sensitivity to gefitinib or erlotinib in non-small cell lung cancer (NSCLC). We investigated the relationships between the two most common types of somatic EGFR mutations, exon 19 deletions and L858R mutations, and clinical outcomes of Korean NSCLC patients after treatment with gefitinib or erlotinib. In Korean patients with NSCLC, EGFR exon 19 deletions and EGFR L858R mutation were identified from tumor specimens obtained before treatment with gefitinib or erlotinib. The response rates, progression-free survival (PFS), and overall survival (OS) were compared between the two groups. A total of 77 patients with either an exon 19 deletion (n = 58) or L858R mutation (n = 19) were treated with gefitinib or erlotinib. The overall response rate was 69%. Patients with an exon 19 deletion had a significantly longer PFS compared with patients with L858R mutation (9.5 vs. 7.7 months; P = 0.029). The L858R mutation was independently associated with a shorter PFS compared with an exon 19 deletion, even after adjusting for other clinical factors (hazard ratio 2.72; 95% CI 1.38-5.38). However, there were no significant differences in response rate (71 vs. 63%) and OS (21.4 vs. 30.7 months) between subjects with exon 19 deletions and L858R mutations, respectively. In Korean NSCLC patients, EGFR exon 19 deletions are associated with longer PFS compared with EGFR L858R mutations. These observations need to be confirmed by large-scale studies of patients.

Abstract 2217: Analysis of mitochondrial variations in non-small cell lung cancer (NSCLC) by microarray-based genome resequencing

Abstract The human mitochondrial genome is a circular double-stranded DNA molecule (mtDNA) of 16,569 base pairs, encoding 13 proteins that are essential in oxidative phosphorylation, two ribosomal genes, and 22 tRNAs. The mtDNA genome has a high copy number per cell, replicates independently of the nuclear genome, and has a maternal-inherited pattern. Mutations in mtDNA genome have long been suspected to play an important role in the development of cancer. Although most cancer cells harbor mtDNA mutations, the question of whether such mutations are associated with clinical prognosis of cancer remains unclear. In this study, we resequenced the entire mitochondrial genomes of tumor tissue from a population of 249 Korean non-small cell lung cancer (NSCLC) patients using the Affymetrix GeneChips Human Mitochondrial Resequencing Array 2.0 (Santa Clara, CA). A total of 892 single nucleotide polymorphisms (SNPs) were identified, including 269 (30.2%) synonymous SNPs, 142 (15.9%) non-synonymous SNPs, 78 (8.7%) SNPs in non-coding region, 188 SNPs (21.1%) in control region, as well as 215 (24.1%) previously unreported SNPs. Each tumor sample contained at least 6 mitochondrial SNPs, with maximum 41 SNPs. We selected 189 (21.2%) SNPs with frequency &amp;gt;2.5% for subsequent analyses. Single SNP analysis using Cox proportional hazards models identified 19 SNPs with p value &amp;lt; 0.05, but none of them remained significant after adjustment for multiple comparisons. Unsupervised clustering analysis revealed two major SNP signature panels, including Signature Panel 1 (8 SNPs) and Signature Panel 2 (5 SNPs). Accordingly, the patient population was divided into three groups: 68 (27.3%) subjects with both signature panels, 73 (29.3%) subjects with only the Signature Panel 1 but not the Signature Panel 2, and 108 (43.4%) subjects without signature panel. In early stage (stage I/ II) NSCLC, patients with the Signature Panel 2 had the worst clinical prognosis, and patients without any signature panel performed best in the survival analyses (adjust hazard ratio=1.83; 95% confidence interval, 1.25-2.66; p=0.003). Further mitochondrial haplogroup analysis identified that the Signature Panel 1 had most defining mutations for haplogroup M and N, and the Signature Panel 2 contained all defining mutations of haplogroup D4a. Interestingly, haplogroup D4a was previous reported as a marker for extreme longevity in Japanese. We conclude that mtDNA mutations may play an important role in survival among early stage lung cancer patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2217.

Dose effect of cigarette smoking on frequency and spectrum of epidermal growth factor receptor gene mutations in Korean patients with non-small cell lung cancer

This study aimed to determine the dose effect of smoking on the mutational frequency and spectrum of epidermal growth factor receptor (EGFR) gene in Korean non-small cell lung cancer (NSCLC). Detailed smoking histories were obtained from 324 consecutively enrolled Korean NSCLC patients. Mutational status of EGFR (exon 18-21) was determined using nested polymerase chain reaction amplification. A total of 108 EGFR mutations (33.3%) were identified in 107 patients. Decreased EGFR mutation rate with increased smoking dose was observed, with 48.0% (82 of 171) in never smokers, 23.1% (15 of 65) in former smokers, and 11.4% (10 of 88) in current smokers. The incidence of EGFR mutation was significantly lower in patients who smoked for more than 25 pack-years (P < 0.0001) or who stopped smoking cigarettes less than 10 years ago (P < 0.0001). Mutations in exon 19 or 21 were associated with fewer total smoke years (5.0 vs. 25.0 years in exon 20, P = 0.024), fewer total pack-years (6.3 vs. 38.9 pack-years in exon 20, P = 0.079), and more smoke-free years (11.1 vs. 3.6 years in exon 20, P = 0.027), compared with those in exon 20. Mutations in exon 19 or 21 were associated with female (P < 0.0001), never smoker (P < 0.0001), and adenocarcinoma (P < 0.0001), whereas those in exon 20 were not. Smoking dosage affects the incidence of EGFR mutations. EGFR mutations in exon 19 or 21 are associated with low exposure to cigarette smoke, whereas EGFR mutation in exon 20 is more common in smokers.